1. Field of the Invention
The present invention relates to a recombinant plasmid for providing plants with resistance against virus having a coding sequence fragment of Papaya ringspot virus helper-component protease gene (PRSV HC-Pro gene). The present invention also relates to a recombinant microorganism. The present invention also relates to a method for providing plants with resistance against virus. The present invention also relates to use of full length or fragment of helper-component protease in manufacturing a plant with resistance to virus.
2. Description of the Prior Arts
Carica papaya L. is one of the high value crops. Effective cures for its virus infection have not been established. Based on progress of plant tissue culture techniques, breakthrough of plant transforming techniques and the concept of pathogen-derived resistance, transferring a portion of viral genome into chromosome of a plant host recently has become a popular means for providing plants with resistance against virus. Transgenic papaya lines were obtained by transforming papaya with the CP gene of PRSV Hawaii strain by microprojectile bombardment (Fitch et al., 1992), but they are not resistant to Taiwan PRSV YK isolate, due to sequence homology-dependent resistance. This limits the application of those transgenic papaya lines in other geographic regions.
Transgenic papaya lines carrying the CP gene of Taiwan PRSV YK isolate were generated by Agrobacterium-mediated transformation (Cheng et al., 1996). Through inoculation of virus and evaluation, the transgenic papaya lines were proven as conferring high level of broad spectrum resistance to PRSV YK as well as distinct PRSV strains from various geographical regions including Hawaii (HA), Mexico (MX) and Thailand (TH) (Bau et al., 2003). YK coat protein transgenic papaya lines confer high levels of resistance to PRSV (Bau et al., 2004). Resistance of transgenic plants carrying coat protein transgene is mediated by the mechanism of post-transcriptional gene silencing (PTGS), PRSV RNA genome is degraded, resulting in achieving resistance against virus. However, the conventional means for providing viral resistance to crops is based on sequence homology. Therefore, a common problem of the conventional means resides in the resistance is virus strain specific (Tennant et al., 2001). Conventional transgenic papaya lines carrying coat protein gene from various virus strains have limited applications in certain geographical regions.
Applicants have generated PRSV YK CP transgenic lines with different levels of resistance to Taiwan PRSV YK strain by introducing coat protein of PRSV YK into premature embryo through Agrobacterium-mediated transformation (Cheng et al., 1996) and selecting out lines with high levels of resistance to virus from various geographical regions such as Hawaii (HA), Thailand (TH) and Mexico (MX) (Bau et al., 2003). The obtained transgenic lines carrying PRSV YK CP gene are subjected to field trials and proven having high levels of resistance to PRSV (Bau et al., 2004).
During the field trials, transgenic resistance of PRSV YK CP transgenic papaya lines is found to be broken down by Papaya ringspot virus super strain. Breakdown of the resistance mediated by coat protein gene is suggested to be caused by helper-component protease (HC-Pro). HC-Pro is known as a gene silencing suppressor, capable of repressing post-transcriptional gene silencing (Anadalakshmi et al., 1998; Brigneti et al., 1998; Shi et al., 1997). Applicants suggest that PRSV 5-19 HC-Pro can suppress gene-silencing mechanism of PRSV YK CP transgenic papaya line. According to failure of using coat protein transgene alone in protecting plants from attacks by super virus strain, a new strategy is developed by the applicants.